title: Real-time imaging of Leishmania mexicana-infected early phagosomes: a study using primary macrophages generated from green fluorescent protein-Rab5 transgenic mice.
abstract: The small GTPase Rab5 is a key regulator of endosome/phagosome maturation and in intravesicular infections marks a phagosome stage at which decisions over pathogen replication or destruction are integrated. It is currently unclear whether Leishmania-infected phagosomes uniformly pass through a Rab5(+) stage on their intracellular path to compartments with late endosomal/early lysosomal characteristics. Differences in routes and final compartments could have consequences for accessibility to antileishmanial drugs. Here, we generated a unique gfp-rab5 transgenic mouse model to visualize Rab5 recruitment to early parasite-containing phagosomes in primary host cells. Using real-time fluorescence imaging of phagosomes carrying Leishmania mexicana, we determined that parasite-infested phagosomes follow a uniform sequence of transient Rab5 recruitment. Residence in Rab5(+) compartments was much shorter compared with phagosomes harboring latex beads. Furthermore, a comparative analysis of parasite life-cycle stages and mutants deficient in lpg1, the gene encoding the enzyme required for synthesis of the dominant surface lipophosphoglycan, indicated that parasite surface ligands and host cell receptors modulate pathogen residence times in Rab5(+) phagosomes, but, as far as tested, had no significant effect on intracellular L. mexicana survival or replication.
authors.affiliation: Department of Molecular Biology, Max-Planck-Institute for Infection Biology, Berlin, Germany.
authors.forename: Christoph
authors.initials: C
authors.lastname: Lippuner
authors.forename: Daniel
authors.initials: D
authors.lastname: Paape
authors.forename: Athina
authors.initials: A
authors.lastname: Paterou
authors.forename: Janko
authors.initials: J
authors.lastname: Brand
authors.forename: Melville
authors.initials: M
authors.lastname: Richardson
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Smith
authors.forename: Kirstin
authors.initials: K
authors.lastname: Hoffmann
authors.forename: Volker
authors.initials: V
authors.lastname: Brinkmann
authors.forename: Clare
authors.initials: C
authors.lastname: Blackburn
authors.forename: Toni
authors.initials: T
authors.lastname: Aebischer
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D005260:Female; D049452:Green Fluorescent Proteins / Q000378:metabolism; D007700:Kinetics; D007894:Leishmania mexicana / Q000502:physiology*; D008024:Ligands; D008264:Macrophages / Q000378:metabolism*; D051379:Mice; D008822:Mice, Transgenic; D010588:Phagosomes / Q000378:metabolism*; D013997:Time Factors; D019076:Transgenes / Q000235:genetics; D020696:rab5 GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D008024:Ligands; D049452:Green Fluorescent Proteins; D020696:rab5 GTP-Binding Proteins